RRC ID 45589
Author Law W, Wuescher LM, Ortega A, Hapiak VM, Komuniecki PR, Komuniecki R.
Title Heterologous Expression in Remodeled C. elegans: A Platform for Monoaminergic Agonist Identification and Anthelmintic Screening.
Journal PLoS Pathog.
Abstract Monoamines, such as 5-HT and tyramine (TA), paralyze both free-living and parasitic nematodes when applied exogenously and serotonergic agonists have been used to clear Haemonchus contortus infections in vivo. Since nematode cell lines are not available and animal screening options are limited, we have developed a screening platform to identify monoamine receptor agonists. Key receptors were expressed heterologously in chimeric, genetically-engineered Caenorhabditis elegans, at sites likely to yield robust phenotypes upon agonist stimulation. This approach potentially preserves the unique pharmacologies of the receptors, while including nematode-specific accessory proteins and the nematode cuticle. Importantly, the sensitivity of monoamine-dependent paralysis could be increased dramatically by hypotonic incubation or the use of bus mutants with increased cuticular permeabilities. We have demonstrated that the monoamine-dependent inhibition of key interneurons, cholinergic motor neurons or body wall muscle inhibited locomotion and caused paralysis. Specifically, 5-HT paralyzed C. elegans 5-HT receptor null animals expressing either nematode, insect or human orthologues of a key Gαo-coupled 5-HT1-like receptor in the cholinergic motor neurons. Importantly, 8-OH-DPAT and PAPP, 5-HT receptor agonists, differentially paralyzed the transgenic animals, with 8-OH-DPAT paralyzing mutant animals expressing the human receptor at concentrations well below those affecting its C. elegans or insect orthologues. Similarly, 5-HT and TA paralyzed C. elegans 5-HT or TA receptor null animals, respectively, expressing either C. elegans or H. contortus 5-HT or TA-gated Cl- channels in either C. elegans cholinergic motor neurons or body wall muscles. Together, these data suggest that this heterologous, ectopic expression screening approach will be useful for the identification of agonists for key monoamine receptors from parasites and could have broad application for the identification of ligands for a host of potential anthelmintic targets.
Volume 11(4)
Pages e1004794
Published 2015-4
DOI 10.1371/journal.ppat.1004794
PMID 25928899
PMC PMC4415803
MeSH Animals Animals, Genetically Modified / genetics Animals, Genetically Modified / metabolism* Anthelmintics / pharmacology* Behavior, Animal / drug effects* Caenorhabditis elegans / drug effects* Caenorhabditis elegans / genetics Caenorhabditis elegans / metabolism Caenorhabditis elegans Proteins / agonists Caenorhabditis elegans Proteins / genetics Caenorhabditis elegans Proteins / metabolism Chloride Channel Agonists / pharmacology* Drosophila Proteins / agonists Drosophila Proteins / genetics Drosophila Proteins / metabolism Drosophila melanogaster Drug Discovery / methods* GTP-Binding Protein alpha Subunits / chemistry GTP-Binding Protein alpha Subunits / genetics GTP-Binding Protein alpha Subunits / metabolism Haemonchus Helminth Proteins / agonists Helminth Proteins / genetics Helminth Proteins / metabolism Humans Hypotonic Solutions / toxicity Interneurons / drug effects Interneurons / metabolism Motor Activity / drug effects Nerve Tissue Proteins / agonists Nerve Tissue Proteins / genetics Nerve Tissue Proteins / metabolism Receptors, Biogenic Amine / agonists Receptors, Biogenic Amine / genetics Receptors, Biogenic Amine / metabolism Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / metabolism Serotonin 5-HT1 Receptor Agonists / pharmacology*
IF 6.158
Times Cited 6
C.elegans tm2654 tm1325 tm2913 tm1846